Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system

Bernd Zetsche; Jonathan S Gootenberg; Omar O Abudayyeh; Ian M Slaymaker; Kira S Makarova; Patrick Essletzbichler; Sara E Volz; Julia Joung; John van der Oost; Aviv Regev; Eugene V Koonin; Feng Zhang

doi:10.1016/j.cell.2015.09.038

Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system

Cell. 2015 Oct 22;163(3):759-71. doi: 10.1016/j.cell.2015.09.038. Epub 2015 Sep 25.

Authors

Affiliations

¹ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Developmental Pathology, Institute of Pathology, Bonn Medical School, Sigmund Freud Street 25, 53127 Bonn, Germany.
² Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
³ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
⁴ National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894, USA.
⁵ Laboratory of Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Dreijenplein 10, 6703 HB Wageningen, Netherlands.
⁶ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Department of Biology, Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
⁷ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Electronic address: zhang@broadinstitute.org.

Abstract

The microbial adaptive immune system CRISPR mediates defense against foreign genetic elements through two classes of RNA-guided nuclease effectors. Class 1 effectors utilize multi-protein complexes, whereas class 2 effectors rely on single-component effector proteins such as the well-characterized Cas9. Here, we report characterization of Cpf1, a putative class 2 CRISPR effector. We demonstrate that Cpf1 mediates robust DNA interference with features distinct from Cas9. Cpf1 is a single RNA-guided endonuclease lacking tracrRNA, and it utilizes a T-rich protospacer-adjacent motif. Moreover, Cpf1 cleaves DNA via a staggered DNA double-stranded break. Out of 16 Cpf1-family proteins, we identified two candidate enzymes from Acidaminococcus and Lachnospiraceae, with efficient genome-editing activity in human cells. Identifying this mechanism of interference broadens our understanding of CRISPR-Cas systems and advances their genome editing applications.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Amino Acid Sequence
CRISPR-Cas Systems*
Endonucleases / chemistry
Endonucleases / genetics*
Francisella / enzymology
Francisella / genetics*
Genetic Engineering / methods*
HEK293 Cells
Humans
Molecular Sequence Data
Nucleic Acid Conformation
RNA, Guide, CRISPR-Cas Systems / genetics
Sequence Alignment

Substances

RNA, Guide, CRISPR-Cas Systems
Endonucleases

Abstract

Publication types

MeSH terms

Substances

Grants and funding